INVARIANT U2 RNA SEQUENCES BORDERING THE BRANCHPOINT RECOGNITION REGION ARE ESSENTIAL FOR INTERACTION WITH YEAST SF3A AND SF3B SUBUNITS

U2 small nuclear RNA (snRNA) contains a sequence (GUAGUA) that pairs w ith the intron branchpoint during splicing. This sequence is contained within a longer invariant sequence of unknown secondary structure and function that extends between U2 stem I and stem IIa. A part bf this region has been proposed to pair with U6 in a structure called helix I II. We made mutations to test the function of these nucleotides in yea st U2 snRNA. Most single base changes cause no obvious growth defects; however, several single and double mutations are lethal or conditiona l lethal and cause a block before the first step of splicing. We used U6 compensatory mutations to assess the contribution of helix III and found that if it forms, helix III is dispensable for splicing in Sacch aromyces cerevisiae. On the other hand, mutations in known protein com ponents of the splicing apparatus suppress or enhance the phenotypes o f mutations within the invariant sequence that connect the branchpoint recognition sequence to stem IIa. Lethal mutations in the region are suppressed by Cus1-54p, a mutant yeast splicing factor homologous to a mammalian SF3b subunit. Synthetic lethal interactions show that this region collaborates with the DEAD-box protein Prp5p and the yeast SF3a subunits Prp9p, Prp11p, and Prp21p. Together, the data show that the highly conserved RNA element downstream of the branchpoint recognition sequence of U2 snRNA in yeast cells functions primarily with the prot eins that make up SF3 rather than with U6 snRNA.